Adventitious root formation and xylogenesis in Arabidopsis thaliana: new insights

The role of indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA) and of the auxin-interacting phytohormone ethylene on xylogenesis is still little known, even if a xylogenic promotion by auxins has been reported. In particular, auxin/ethylene-target tissue(s), modality of the de novo xylary process, and the kind of ectopic elements formed (metaxylem vs. protoxylem) are currently unknown. It is instead widely known that auxins positively affect adventitious root (AR) formation, e.g. in the model plant Arabidopsis thaliana and in in vitro cultured systems of numerous species. It is also unclear whether IBA may act on the xylogenic process and AR formation independently of its conversion into IAA. To investigate these topics, histological analyses were carried out in the hypocotyls of Arabidopsis thaliana wild type seedlings and ech2ibr10 and ein3eil1 mutants, which are blocked in IBA-to-IAA conversion and ethylene signalling, respectively. Moreover, in Arabidopsis in planta, both AR formation and xylogenesis are known to start from pericycle founder cells of the hypocotyl basal part (Della Rovere et al., Annals of Botany, 2015). The thin cell layer (TCL) system has been useful for investigating AR formation and xylogenesis out of the plant context, but in the presence of exogenous auxin combined with kinetin, with both programs starting from stem endodermis derivatives. It is unknown whether Arabidopsis TCLs produce ARs and/or xylary elements under IBA alone (10 μM) or IAA alone (10 μM), and whether they contain endogenous IAA/IBA at culture onset. Moreover, it is unknown whether an IBA-to-IAA conversion is also active in TCLs, and positively affects ARs/xylary cells formation, with/without ethylene signalling involvement. Results. In planta, IAA favoured protoxylem formation, whereas IBA induced ectopic metaxylem formation with ethylene cooperation through the EIN3EIL1 network. In TCLs, undetectable levels of both auxins were present at culture onset, and ARs were better induced by IBA alone than IAA alone, with an inverse relationship with xylogenesis at the same exogenous concentration. The AR response of IBA-treated TCLs from ech2ibr10 and ein3eil1 mutants was strongly reduced. Nitric oxide (NO), an IAA downstream signal and a by-product of IBA-to-IAA conversion, was also early detected in IAA- and IBA-treated TCLs, but at higher levels in the latter explants, showing that IBA-induced AR formation by conversion into IAA involves NO activity. All together results show that IBA to IAA conversion is important for AR formation, and reveal that the cross-talk between auxin and ethylene is important for AR formation and xylogenesis, occurring through the EIN3/EIL1 signalling pathway.